RESUMO
Chaperone-mediated autophagy (CMA) serves as a critical upstream regulator of lipophagy and lipid metabolism in hepatocyte. However, the role of CMA in lipid metabolism of macrophage, the typical component of atherosclerotic plaque, remains unclear. In our study, LAMP-2A (L2A, a CMA marker) was reduced in macrophages exposed to high dose of oleate, and lipophagy was impaired in advanced atherosclerosis in ApoE (-/-) mice. Primary peritoneal macrophages isolated from macrophage-specific L2A-deficient mice exhibited pronounced intracellular lipid accumulation. Lipid regulatory enzymes, including long-chain-fatty-acid-CoA ligase 1 (ACSL1) and lysosomal acid lipase (LAL), were increased and reduced in L2A-KO macrophage, respectively. Other lipid-related proteins, such as SR-A, SR-B (CD36), ABCA1, or PLIN2, were not associated with increased lipid content in L2A-KO macrophage. In conclusion, deficient CMA promotes lipid accumulation in macrophage probably by regulating enzymes involved in lipid metabolism. CMA may represent a novel therapeutic target to alleviate atherosclerosis by promoting lipid metabolism. Graphical abstract.
Assuntos
Aterosclerose/metabolismo , Autofagia , Metabolismo dos Lipídeos , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Macrófagos Peritoneais/metabolismo , Animais , Aterosclerose/genética , Aterosclerose/patologia , Autofagia/efeitos dos fármacos , Células Cultivadas , Coenzima A Ligases/metabolismo , Modelos Animais de Doenças , Metabolismo dos Lipídeos/efeitos dos fármacos , Proteína 2 de Membrana Associada ao Lisossomo/genética , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Ácido Oleico/toxicidade , Esterol Esterase/metabolismoRESUMO
Chaperone-mediated autophagy (CMA) is one of the three types of autophagy. In recent years, CMA has been shown to be associated with the pathogenesis of several types of cancer. However, whether CMA is involved in the pathogenesis of colorectal cancer (CRC) remains unclear. In this study, we investigated CMA activity in tissue specimens from CRC patients and mouse models of colitis-associated CRC (induced by administration of AOM plus DSS). In addition, we down-regulated CMA in CT26 colon carcinoma cells stably transfected with a vector expressing a siRNA targeting LAMP-2A, the limiting component in the CMA pathway, to explore the role of CMA in these cells. Apoptosis was detected using TUNEL assay, and the apoptosis-related proteins were detected using western blotting. Cell proliferation was assessed using MTT assay, Ki-67 labelling and western blotting for PCNA. We found that LAMP-2A expression was significantly increased in CRC patients and mouse models and varied according to the stage of the disease. Inhibition of CMA in CT26 cells facilitated apoptosis, as evidenced by increased TUNEL immunolabeling, increased expression of Bax and Bnip3, and decreased expression of Bcl-2. Cell proliferation assays showed that inhibition of CMA impeded the proliferation of CT26 cells. These data support the hypothesis that CMA is up-regulated in CRC, and inhibition of CMA may be a new therapeutic strategy for CRC patients.